The oral cavity supports a rich and diverse microbial population. Oral health is dependent on the maintenance of stable microbial communities; disease occurs when this balance is disturbed and more pathogenic species outgrow the commensals. Health and disease in the mouth are active processes in which the ecology of communities, not of single organisms, is paramount.

Expert authors from around the world provide an update on recent developments in the burgeoning field of oral microbial ecology. The focus of the book is on the most topical areas in oral microbiology and the volume is a major new work in the field. The chapters are arranged into five sections: microbial populations in oral biofilms, the structure of oral biofilms, communication and sensing within biofilms, health to disease - the microbial community perspective, and new approaches for oral biofilm control. Specialist authors contribute chapters on various topics including population biology, detection and culture of novel oral bacteria, bacterial catabolism of salivary substrates, structural organization of oral biofilms, the extracellular polysaccharides matrix, extracellular proteins and DNA in the matrix, a holistic view of inter-species bacterial interactions, environmental sensory perception, microbial community interactions of Streptococcus mutans, biofilms in periodontal health and disease, oral biofilms as a reservoir for pathogens, oral biofilms as a device for therapeutic agents, and probiotics in oral healthcare.

The book is an essential text for scientists interested in oral microbiology, bacterial communities and biofilms and is recommended reading for anyone working in the areas of oral health, and the pathogenesis of dental caries and periodontal disease. A recommended book for all microbiology laboratories.

In this chapter we consider the biology of the viridans streptococci in the human oropharynx with a particular focus on the pioneer bacterium Streptococcus mitis. We show that, although this species is a constant component of the human oral cavity, each person harbors a unique and diverse population of stains that appear not to be shared within a family and, apparently, are rarely transmitted from mother to neonate. The population of stains of S. mitis within the mouth of each individual exhibits turnover perhaps in response to pressure exerted by the mucosal immune system since it has been shown that some secretory immunoglobulin A (SIgA) antibodies are clone-specific. We assert that the strains that are successful in establishing in the mouth are physiologically adapted to occupy their niche within their habitat. While it is clear that in vitro experiments and animal models have provided useful information, they are no substitute for studying commensal oral bacteria in their environment, the human oral cavity.

2. Detection and Culture of Novel Oral Bacteria

William Wade

The oral microbiota is highly diverse and includes fungi, protozoa, viruses and bacteria. Both domains of prokaryotes, Archaea and Bacteria are present. Representatives of the Archaea are restricted to a few taxa in the genus Methanobrevibacter, while there are over 600 species of Bacteria, from at least 12 phyla. The full diversity of bacterial populations in the mouth has been recognised following the application of culture-independent methods of analysis, based on 16S rRNA gene sequence comparisons. Because oral bacteria are typically slow-growing and fastidious, and around half cannot be grown in the laboratory at all, the taxonomic process of classifying and naming bacterial species is ongoing and over 100 cultivable taxa have still to be named. In recent years, attempts have been made to culture the not-yet-cultured portion of the microbiota. There are a number of reasons why certain taxa are uncultivable and these include a need for a specific nutrient, extreme oxygen sensitivity and dependence on other organisms. The inter-dependence among members of the oral microbial community may relate to cooperative degradation of natural substrates for growth or the need to participate in signalling networks that control growth rate and resuscitation from dormancy. Novel culture media and methods are being developed that reproduce the in vivo environment and thus encourage previously uncultured organisms to grow in the laboratory.

3. Bacterial Catabolism of Salivary Substrates

David Beighton, Sadaf Rasheed Mughal and Thuy Do

The oral biofilm proliferates in the mouth by primarily utilizing components of saliva as dietary foods are rapidly cleared. The complex microbial community functions in a concerted manner to obtain nutrients, sugars and amino acids, from salivary components including mucins, by the production of a range of glycosidic enzymes including sialidase, β-galactosidase, N-acetylglucosaminidases, α-fucosidase and mannosidases and exo- and endo-proteolytic activities. Degradation of glycans occurs sequentially and in vitro studies indicate that liberated sugars are rapidly transported though evidence of cross-feeding between species, utilizing liberated sugars, is evident. Streptococcus oralis is a species with the greatest ability to deglycosylate both N- and O-linked glycans and has been used extensively in model systems. New research should take advantage of modern high throughput sequencing techniques to determine the biofilm transcriptome of humans receiving defined diet, including fasting, to ascertain the response of the biofilm to in vivo conditions.

The more we learn about the oral microbiota, the more we have to accept its high degree of complexity and diversity. The composition of the oral microbial community depends on a number of patient-specific factors; the immunology and genetic background of patient, underlying diseases (chronic or acute, e.g. diabetes, infections), hormone status (e.g. pregnancy), medication (e.g. immunosuppressive therapy or antibiotics) and behaviour of the patient (e.g. nutrition, oral hygiene, smoking habits). It even differs within the oral cavity of a subject due to the ecological niches influenced by different variables such as oxygen level, nutrition, physical stress and saliva flow. All these factors influence the combination and multitude of bacterial species that are finally responsible for maintaining health or causing disease within the oral ecosystem. However, bacteria do not function as isolated entities but live together in complex sessile communities called oral biofilms. The efficiency of oral biofilms depends on coordinated, metabolically integrated, spatially organized networks. Therefore, it is crucial to understand these highly dynamic communities and to observe and analyze their composition and architecture. In this chapter we first discuss the methodology used in oral biofilm structure research and subsequently discuss the current status of the research as well as our concept of supra- and subgingival biofilm organization.

5. The Role of Extracellular Polysaccharides Matrix in Virulent Oral Biofilms

Virulent biofilms are responsible for a range of infections, including those occurring in the mouth. The disease dental caries is a prime example of the consequences arising from interactions of host, bacteria and diet (sugars), resulting in the assembly of pathogenic biofilms on susceptible tooth-surfaces. All biofilms harbor a microbial-derived extracellular-matrix. The exopolysaccharides (EPS) formed on the tooth-pellicle and on the bacterial surfaces provide binding sites for microorganisms; eventually the accumulated EPS enmeshes the microbial cells. The metabolic activity of the bacteria embedded in EPS-rich and diffusion-limited matrix leads to acidification of the milieu, and subsequent onset of the disease through acid dissolution of enamel (expressed clinically as cavities). This chapter focuses on the mechanisms through which the EPS produced by Streptococcus mutans modulates the assembly of a spatially complex 3D matrix during biofilm morphogenesis within a mixed-bacterial species community. The architecture of the matrix influences the development of pH heterogeneity in the 3D environment of intact biofilms. The matrix facilitates the formation of structured acidic-microenvironments in close proximity to the apatite-surface, which are essential for the evolution of virulent biofilms formed on the tooth. These observations may have relevance beyond the mouth, as matrix is inherent to all biofilms.

6. Extracellular Proteins and DNA in the Matrix of Oral Biofilms

Nicholas S. Jakubovics

A core component of microbial biofilms is the extracellular matrix, which binds together the assembled micro-organisms and regulates the passage of small molecules to and from cells. The scaffolding of the matrix is composed of biological macromolecules including carbohydrates, nucleic acids and proteins. The production and function of extracellular polysaccharides in oral biofilms has been the subject of a great deal of research over many decades, and is considered in a separate chapter. More recently, it has become clear that proteins and extracellular DNA (eDNA) play key roles in maintaining the structure of many different biofilms, including oral biofilms. This chapter reviews the recent research on proteins and eDNA in the biofilm matrix, and discusses the evidence that oral micro-organisms utilise these macromolecules for specific functions within mixed-species biofilms.

Mature dental-plaque biofilm communities contain hundreds of bacterial species. The potential for these communities to cause caries or periodontal disease relates to bacterial spatiotemporal biofilm development and species composition. At least three forms of inter-species interactions can conceivably mediate altered biofilm development and species composition. These are coaggregation, metabolic interactions, and cell-cell signaling. Coaggregation is the specific recognition and adhesion of different species of bacteria and likely contributes toward the ordered (sequential) integration of species into biofilms as well as improving species retention in a flowing environment. 'Metabolic interactions' is an umbrella term that describes the exchange of metabolites or environmental protection afforded between adjacent species within dental plaque. Cell-cell signaling is a phenomenon that has gained increasing research interest over the past decade. One broad inter-species signaling molecule system consists of a collection of inter-convertible cell-cell signal molecules that are collectively called autoinducer-2 (AI-2). Evidence indicates that AI-2 can alter bacterial phenotypes, when present in saliva at concentrations as low as the nanomolar range. It is the aim of this chapter to describe each of these inter-species phenomena, with case-examples, and extrapolate singular and combined roles in the spatio-temporal development of dental plaque. The potential for these phenomena to create shifts in community species composition have implications for the development of polymicrobial diseases.

8. Environmental Sensory Perception by Oral Streptococci

Justin Merritt and Jens Kreth

Oral streptococci encounter an exceptionally wide range of environmental stresses and population densities. These stimuli are sensed by efficient detection systems that also coordinate the appropriate adaptive genetic responses. The majority of these detection systems utilize membrane bound sensory proteins that are directly or indirectly regulated by their sensed stimuli. Such systems play an intimate role in mitigating the potential damage caused by changes in redox potential, fluctuations in local pH, and toxicity from antimicrobial agents. In addition, the typical life cycle of oral streptococci includes a transition from growth in a relatively low cell density planktonic state to an extremely high cell density biofilm environment. Consequently, various sensory systems are dedicated to detecting this increase in population density and regulating the genetic pathways that are essential for persistence in a highly competitive multispecies biofilm environment. Recent studies have identified many of the targets of these sensory systems and have provided unprecedented insight into the intimate connection between the constantly changing oral environment and the genetic machinery of oral bacteria.

9. Microbial Community Interactions of the Cariogenic Organism Streptococcus mutans

Saswat Sourav Mohapatra and Indranil Biswas

The human oral cavity is estimated to host more than 700 bacterial species formed into distinct biofilm communities, of which more than 50% are yet to be cultured in the laboratory. Though oral streptococci constitute two thirds of the total commensals, only a fraction known as mutans streptococci are involved in producing dental caries. The oral streptococci are the primary colonizers of the tooth and other mucosal surfaces in the oral cavity and initiate plaque biofilm formation. Mutual interaction in the form of cooperation and competition shapes the constitution of the oral microflora. Colonization by Streptococcus mutans, having significant acidogenicity and aciduricity properties, is primarily responsible for dental caries formation. Recent advances in nucleotide sequencing and other high throughput methods have provided significant clues to the biology and gene regulation of S. mutans in a community structure. Many therapeutic methods are being devised to specifically target the S. mutans in the biofilm, without disturbing other bacterial species. Significant among them are targeting the interbacterial signaling, replacing cariogenic flora with non-cariogenic flora, and specifically targeted antimicrobial peptides (STAMPs). As the research progresses in this field, better therapeutic methods are on the horizon.

10. Biofilms in Periodontal Health and Disease

Purnima S Kumar, Matthew R Mason and Janel Yu

Dental plaque biofilm is composed of a diverse microbial community. Several decades of research have been focused on the role played by the subgingival biofilm in the etiology of periodontal diseases. However, recent evidence from other ecosystems within the human body indicates that these biofilms also play an important role in maintaining health. The purpose of this chapter is to explore the health benefits of subgingival plaque, and to outline the development of the biofilm as well as to characterize the bacteria present in the plaque biofilm in health and disease. Additionally, the virulence mechanisms of health and disease-associated biofilms will be outlined.

11. Periodontal Biofilm and Immunity: Immune Subversion by Select Pathogens as a Community Service

George Hajishengallis

The polymicrobial community that initiates periodontal disease does not represent a random compilation of bacteria. Rather, these bacteria form organized consortia that have evolved through mutually beneficial relationships. This chapter focuses on microbial immune subversion as a means by which select pathogens may contribute to the adaptive fitness of the entire periodontal biofilm. For instance, Porphyromonas gingivalis expresses specialized virulence traits that undermine immunity and promote non-resolving inflammation, which, respectively, protect the bacteria and facilitate nutrient acquisition. The virulence factors involved (e.g. cysteine proteases and atypical lipopolysaccharide structures) are released as components of readily diffusible membrane vesicles, which can thus become available for the benefit of other biofilm organisms. The elucidation of immune subversion mechanisms of key periopathogens that promote the collective virulence of their communities may provide new avenues of therapeutic intervention in human periodontitis.

The link between oral flora and lung infections in mechanically ventilated (MV) patients (the 'Oral Systemic Link') has always been circumstantial, based on clinical and nursing practices and preventative care. At the turn of the 20th century, that began to change as molecular and imaging methods provided tools to better evaluate microbial exchange, refocusing on the importance of the endotracheal tube (ETT) lumen as a potential conduit, devoid of normal cellular defensive components. We undertook the challenge in two phases, first engineering the Adult -Ventilator Endotracheal Lung (A-VEL) simulator to replicate the closed, bi-directional airway and stress of the intubated patient in the intensive care unit (ICU). Of singular importance, was the incorporation of multiple quantitative imaging techniques to define the 3-D biofilm luminal development in Stages (I-IV) from single to complex microbial communities and the incorporation of dental pathogens (Streptococcus mutans and Porphyromonas gingivalis) as an endogenous event, in the preconditioning of the ETT luminal surface, an abiotic medical device. The second phase shifted to the in vivo environment and in multiple clinical studies, unmasked the bi-phasic nature of ETT luminal colonization, oral-endogenous (Early) to systemic - exogenous (Late) at a 3-5 day 'switch'. Further, the ICU studies dramatized the shift from the infectious process in VAP to Work of Breathing (WOB), where the former occurred in 16%, while biofilm accretion, occlusion of the ETT lumen and increased airway resistance occurred in 100% of intubated patients. Most recently, we have used both 16S (microarray) and 18S rRNA (pyrosequencing) to redefine the proportions of bacteria and fungi from oral reservoirs, and been astonished by the richness and diversity of the oral fungal community in the ETT accretion occlusion, often yielding >15 species . Endotracheal Tube Associated Pneumonia (EAP) management continues to elude optimal strategies, but the use of selected oral probiotics coupled with better oral care in both the ICU and admitting institutions is gaining reinforcement: "Oral Stewardship". Further, the utilization of dental professionals in the ICU has importance, as has the recognition that the next fertile area of airway disease (oral to systemic) study is the neonatal intensive care unit (NICU), where 50% of newborns may be intubated and develop EAP with no teeth. How? Why?

An overview is presented on oral biofilm formation and recent developments in oral biofilm control using mechanical devices (manual or powered toothbrushes and interdental cleaning devices) and biofilm control based on oral chemotherapeutics (antibacterial toothpastes and mouthrinses). For clinical efficacy of oral chemotherapeutics, it is important that the antibacterial remains active in the oral cavity for periods longer than the actual brushing or rinsing time, a characteristic called 'substantivity'. Substantivity can be achieved by adsorption of antibacterials to oral hard and soft tissues followed by release. Mechanical cleaning never results in complete removal of oral biofilm: most notably in fissures, interproximal spaces, gingival pockets and around orthodontic appliances. Recently, it has been demonstrated that this residual biofilm can act as a reservoir for oral chemotherapeutics that are slowly released over time in bio-active concentrations. This function of oral biofilm was already known for fluoride and has been demonstrated to aid in preventing caries. Results from our laboratory showed that residual biofilm after mechanical cleaning can release absorbed antibacterial agents from toothpastes and mouthrinses in bio-active concentrations.

14. Probiotics: a Possible Tool in Oral Health Care?

Christof Godts, Gitte Loozen, Marc Quirynen and Wim Teughels

The human oral cavity is colonized by a wide variety of bacteria, which form very complex and dynamic biofilms on hard and soft tissues. Certain members of these microbiological communities are associated with oral infections, such as caries and periodontal diseases. New treatment approaches are emerging that do not rely on conventional antibiotic therapies, since complete eradication of pathogenic bacteria from oral biofilms is impossible and antibiotic resistance is becoming problematic. For example, attempts have been made to reduce the overall pathogenicity of tissue-associated biofilms by introducing live beneficial bacteria. Early successes, primarily in the field of gastro-intestinal microbiology, have paved the way for the introduction of probiotics in oral health care. These new anti-/pro-microbial therapies are considered very promising for prevention and treatment of plaque related oral diseases. In this chapter, the concept of probiotics for oral healthcare is introduced, followed by an overview of the diverse mechanisms of probiotic action in the oral cavity. Since the benefits of probiotics will ultimately be revealed by clinical studies, the clinical outcomes of probiotic applications for combating dental caries and periodontal diseases are addressed. Finally the interactions of probiotics with the oral microbial ecosystem are discussed and future perspectives regarding the oral probiotic concept are presented.